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1.
World J Surg Oncol ; 19(1): 108, 2021 Apr 10.
Artigo em Inglês | MEDLINE | ID: mdl-33838675

RESUMO

BACKGROUND: Mastoscopic surgery is proven to have lower incidence of postoperative complications and better postoperative recovery than traditional breast cancer surgery. This study aimed to examine the feasibility of mastoscopic modified radical mastectomy (MRM) with skin nipple-areola preservation under air cavity-free suspension hook and stage I silicone prosthesis implantation (SMALND) compared with routine MRM. METHODS: This was a retrospective study of patients who underwent MRM for breast cancer at the Shengjing Hospital Affiliated to China Medical University between January 1, 2019, and June 30, 2019. Surgical outcomes, complications, satisfaction, and quality of life (Functional Assessment of Cancer Therapy-Breast [FACT-B] [Chinese version]) were compared between the two groups. RESULTS: A total of 87 patients were enrolled, with 30 underwent SMALND and 57 underwent routine MRM. The intraoperative blood loss in the SMALND group was lower than in the control group (165.3±44.1 vs. 201.4±52.7 ml, P=0.001), the operation time was longer (220.5±23.9 vs. 155.6±9.2 min, P<0.001), daily axillary drainage volume was smaller (20.2±3.6 vs. 24.1±3.0 ml, P<0.001), daily subcutaneous drainage volume was smaller (15.5±2.3 vs. 19.3±3.5 ml, P<0.001), the discharge time was shorter (7.5±1.6 vs. 9.0±1.8 days, P<0.001), and FACT-B scores were higher (83.8±5.6 vs. 72.1±4.6, P<0.001). The overall satisfaction was higher in the SMALND group than in the controls (76.7% vs. 54.4%, P=0.041). Compared with the controls, the occurrence rates of nipple and flap necrosis, upper limb edema, and paraesthesia in the SMALND group were lower within 6 months (all P<0.05). CONCLUSIONS: Compared with traditional MRM, SMALND had better surgical outcomes, higher satisfaction, higher quality of life, and lower complication rates.

2.
BMC Med Inform Decis Mak ; 21(1): 86, 2021 03 06.
Artigo em Inglês | MEDLINE | ID: mdl-33676504

RESUMO

BACKGROUND: Colorectal cancer (CRC) is a common malignancy worldwide. Despite being the most common cancer in Singapore, CRC screening rate remains low due to knowledge deficits, social reasons such as inconvenience and a lack of reminder or recommendation. A decision aid (DA) may facilitate an individual's decision-making to undertake CRC screening by addressing misconceptions and barriers. We postulate that a more person-centred and culturally adapted DA will better serve the local population. The views of the target users are thus needed to develop such a DA. A CRC screening DA prototype has been adapted from an American DA to cater to the Asian users. This study aimed to explore user perspectives on an adapted CRC screening DA-prototype in terms of the design, content and perceived utility. METHODS: The study used in-depth interviews (IDIs) and focus group discussions (FGDs) to gather qualitative data from English-literate multi-ethnic Asian adults aged 50 years old and above. They had yet to screen for CRC before they were recruited from a public primary care clinic in Singapore. The interviews were audio-recorded, transcribed and analysed to identify emergent themes via thematic analysis. RESULTS: This study included 27 participants involved in 5 IDI and 5 FGDs. Participants found the DA easily comprehensible and of appropriate length. They appreciated information about the options and proposed having multi-lingual DAs. The design, in terms of the layout, size and font, was well-accepted but there were suggestions to digitalize the DA. Participants felt that the visuals were useful but there were concerns about modesty due to the realism of the illustration. They would use the DA for information-sharing with their family and for discussion with their doctor for decision making. They preferred the doctor's recommendation for CRC screening and initiating the use of the DA. CONCLUSIONS: Participants generally had favourable perceptions of the DA-prototype. A revised DA will be developed based on their feedback. Further input from doctors on the revised DA will be obtained before assessing its effectiveness to increase CRC screening rate in a randomized controlled trial.


Assuntos
Neoplasias Colorretais , Detecção Precoce de Câncer , Adulto , Grupo com Ancestrais do Continente Asiático , Neoplasias Colorretais/diagnóstico , Técnicas de Apoio para a Decisão , Conhecimentos, Atitudes e Prática em Saúde , Humanos , Pessoa de Meia-Idade , Pesquisa Qualitativa , Singapura
3.
Medicine (Baltimore) ; 100(10): e25140, 2021 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-33725916

RESUMO

BACKGROUND: Human immunodeficiency virus (HIV)-associated distal symmetric peripheral neuropathy (DSPN) is one of the most frequent neurological complications of HIV infection, and causes pain and dysaesthesias in millions globally. Many individuals with this infection report using acupuncture to manage their symptoms, but evidence supporting the use of acupuncture is limited. This systematic review will assess the effectiveness and safety of acupuncture for patients with HIV-associated DSPN. METHODS: Databases including MEDLINE, EMBASE, the Cochrane Central Register of Controlled Trials, Scopus, Web of science, AMED (Allied and Complementary Medicine), the Chinese Biomedical Literature Database, the China National Knowledge Infrastructure, Wanfang Database, VIP Database and clinical trials registers (the WHO International Clinical Trials Registry Platform portal and www.ClinicalTrials.gov) will be electronically searched from inception to December 1, 2020. All randomized controlled trials in English or Chinese without restriction on publication status will be included. Selection of studies, extraction of data, and assessment of studies quality will be independently performed by 2 reviewers. The primary outcome measure will be the change in pain intensity assessed by validated scales. Secondary outcomes include change in neurologic summary scores, quality of life, physical function evaluated by admitted tools, and adverse events related to acupuncture reported in the included trials. If possible, a meta-analysis will be conducted to provide an estimate of the pooled treatment effect using Review Manager 5.3 statistical software. Otherwise, qualitative descriptive analysis will be given. The results will be presented as the risk ratio for binary data and the mean difference (MD) or standardized MD for continuous data. RESULTS: The results of the systematic review will be disseminated via publication in a peer-reviewed journal and presented at a relevant conference. CONCLUSION: This review will be the first review entirely focused on assessing the effectiveness and safety of acupuncture for HIV-associated DSPN. PROSPERO REGISTRATION NUMBER: CRD42020210994.


Assuntos
Terapia por Acupuntura/efeitos adversos , Infecções por HIV/complicações , Neuralgia/terapia , Parestesia/terapia , Polineuropatias/terapia , Infecções por HIV/imunologia , Infecções por HIV/virologia , Humanos , Metanálise como Assunto , Neuralgia/diagnóstico , Neuralgia/imunologia , Neuralgia/virologia , Medição da Dor , Parestesia/diagnóstico , Parestesia/imunologia , Parestesia/virologia , Polineuropatias/diagnóstico , Polineuropatias/imunologia , Polineuropatias/virologia , Qualidade de Vida , Ensaios Clínicos Controlados Aleatórios como Assunto , Índice de Gravidade de Doença , Revisões Sistemáticas como Assunto , Resultado do Tratamento
4.
PLoS One ; 16(3): e0248235, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-33667285

RESUMO

PURPOSE: We aim to investigate the risk factors associated with the prognosis of proliferative diabetic retinopathy (PDR) after a sequential treatment of intravitreal injection of bevacizumab (IVB) and pars plana vitrectomy (PPV). METHODS: In this cohort study, 63 eyes from 55 patients (21 females) diagnosed with PDR, who needed PPV for non-clearing vitreous hemorrhage or fibrovascular membrane proliferation were enrolled. All the eyes underwent IVB followed by PPV. Anterior chamber tap was performed at the beginning of both procedures to evaluate the concentration of vascular endothelial growth factor (VEGF), interleukin (IL)-6, IL-8, and monocyte chemoattractant protein (MCP)-1. RESULTS: Forty-seven patients (54 eyes) were followed over six months, averaging 12±5 (6-19) months. The concentration of VEGF significantly decreased after IVB (P<0.001), while other cytokines did not change significantly. The aqueous humor level of IL-8 after IVB (R = 0.378, P = 0.033), MCP-1 before (R = 0.368, P = 0.021) and after (R = 0.368, P = 0.038) IVB, and combined phacoemulsification (R = 0.293, P = 0.032) was correlated with the logMAR visual acuity at the last follow-up. Multivariate analysis showed that MCP-1 was the predictor for a worse visual outcome (B = 0.108, 95% CI 0.013-0.202; P = 0.027). CONCLUSIONS: MCP-1 was a predictor for the unfavorable visual outcome of PDR after IVB pretreatment and PPV.

5.
Artigo em Inglês | MEDLINE | ID: mdl-33719413

RESUMO

As a promising candidate in various fields, including energy conversion and electronics, layered van der Waals metal phosphorus trichalcogenides (MPX3) have been widely explored. In addition to the layered structures, MPX3 comprising post-transition metals (i.e., Sn and Pb) are known to form a unique 3D framework with nonlayered structure. However, the nonlayered two-dimensional (2D) crystals of this family have remained unexplored until now. Herein, we successfully synthesized 2D nonlayered tin thiohypodiphosphate (Sn2P2S6) nanosheets, having an indirect bandgap of 2.25 eV and a thickness down to ∼10 nm. The as-obtained nanosheets demonstrate promising photocatalytic water splitting activity to generate H2 in pure water under simulated solar light (AM 1.5G). Moreover, the ultrathin Sn2P2S6 catalyst shows auspicious performance and stability with a continuous operation of 40 h. This work is not only an expansion of the MPX3 family, but it is also a major milestone in the search for new materials for future energy conversion.

6.
Biochem Biophys Res Commun ; 547: 125-130, 2021 Apr 02.
Artigo em Inglês | MEDLINE | ID: mdl-33610040

RESUMO

ABJECTIVE: Interaction of hypertension and hyperhomocysteinemia (HHcy) leads to enhanced cardiac remodeling in hypertensive heart disease. However, the mechanism of collagen accumulation and cardiac remodeling remains unclear. In this study, we attempted to evaluate the relationship between hypertension and HHcy in the context of cardiac remodeling and to explore its mechanism of action. METHODS: Wistar Kyoto (WKY) and spontaneous hypertension rats (SHR) were randomly divided into four groups, namely WKY group, WKY + HHcy group, SHR group and SHR + HHcy group. We measured blood pressure (BP), plasma homocysteine (Hcy), serum superoxide dismutase (SOD) and serum malondialdehyde (MDA). We also examined cardiac histopathology and gene and protein expression levels of Nrf2 and HO-1. RESULTS: Compared with the WKY group, myocardial interstitial and perivascular collagen deposition in the WKY + HHcy group, the SHR group and the SHR + HHcy group increased successively, indicating that cardiac remodeling gradually increased, and HHcy aggravated cardiac remodeling was more serious in hypertensive rats. SOD decreased gradually in the four groups, while MDA was on the contrary. WKY + HHcy and SHR + HHcy groups both suppressed Nrf2 and HO-1 expression and inhibited the translocation of Nrf2 from cytoplasm to nucleus compared with their control groups, and the SHR + HHcy group had a stronger inhibitory effect. CONCLUSION: HHcy enhanced cardiac remodeling in rats by enhancing oxidative stress, suppressing the Nrf2/HO-1 pathway and Nrf2 nuclear transport, and this inhibitory effect was stronger in the context of hypertension.

8.
ACS Appl Mater Interfaces ; 13(5): 5907-5918, 2021 Feb 10.
Artigo em Inglês | MEDLINE | ID: mdl-33506676

RESUMO

Critical limb ischemia (CLI) is a severe form of peripheral artery disease (PAD). It is featured by degenerated skeletal muscle and poor vascularization. During the development of CLI, the upregulated matrix metalloproteinase-2 (MMP-2) degrades muscle extracellular matrix to initiate the degeneration. Meanwhile, MMP-2 is necessary for blood vessel formation. It is thus hypothesized that appropriate MMP-2 bioactivity in ischemic limbs will not only attenuate muscle degeneration but also promote blood vessel formation. Herein, we developed ischemia-targeting poly(N-isopropylacrylamide)-based nanogels to specifically deliver an MMP-2 inhibitor CTTHWGFTLC (CTT) into ischemic limbs to tailor MMP-2 bioactivity. Besides acting as an MMP-2 inhibitor, CTT promoted endothelial cell migration under conditions mimicking the ischemic limbs. The nanogels were sensitive to the pH of ischemic tissues, allowing them to largely aggregate in the injured area. To help reduce nanogel uptake by macrophages and increase circulation time, the nanogels were cloaked with a platelet membrane. An ischemia-targeting peptide CSTSMLKA (CST) was further conjugated on the platelet membrane for targeted delivery of nanogels into the ischemic area. CTT gradually released from the nanogels for 4 weeks. The nanogels mostly accumulated in the ischemic area for 28 days. The released CTT preserved collagen in the muscle and promoted its regeneration. In addition, CTT stimulated angiogenesis. Four weeks after CLI, the blood flow and vessel density of the ischemic limbs treated with the nanogels were remarkably higher than the control groups without CTT release. These results demonstrate that the developed nanogel-based CTT release system has the potential to stimulate ischemic limb regeneration.


Assuntos
Isquemia/tratamento farmacológico , Metaloproteinase 2 da Matriz/metabolismo , Inibidores de Metaloproteinases de Matriz/farmacologia , Músculo Esquelético/efeitos dos fármacos , Nanogéis/química , Neovascularização Patológica/tratamento farmacológico , Animais , Sistemas de Liberação de Medicamentos , Feminino , Humanos , Isquemia/patologia , Inibidores de Metaloproteinases de Matriz/síntese química , Inibidores de Metaloproteinases de Matriz/química , Camundongos , Camundongos Endogâmicos C57BL , Estrutura Molecular , Músculo Esquelético/patologia , Neovascularização Patológica/patologia , Tamanho da Partícula , Propriedades de Superfície
9.
J Control Release ; 331: 376-389, 2021 Mar 10.
Artigo em Inglês | MEDLINE | ID: mdl-33508351

RESUMO

Stem cell transplantation has been extensively explored to promote ischemic limb vascularization and skeletal muscle regeneration. Yet the therapeutic efficacy is low due to limited cell survival under low oxygen environment of the ischemic limbs. Therefore, continuously oxygenating the transplanted cells has potential to increase their survival. During tissue regeneration, the number of blood vessels are gradually increased, leading to the elevation of tissue oxygen content. Accordingly, less exogenous oxygen is needed for the transplanted cells. Excessive oxygen may induce reactive oxygen species (ROS) formation, causing cell apoptosis. Thus, it is attractive to develop oxygen-release biomaterials that are responsive to the environmental oxygen level. Herein, we developed oxygen-release microspheres whose oxygen release was controlled by oxygen-responsive shell. The shell hydrophilicity and degradation rate decreased as the environmental oxygen level increased, leading to slower oxygen release. The microspheres were capable of directly releasing molecular oxygen, which are safer than those oxygen-release biomaterials that release hydrogen peroxide and rely on its decomposition to form oxygen. The released oxygen significantly enhanced mesenchymal stem cell (MSC) survival without inducing ROS production under hypoxic condition. Co-delivery of MSCs and microspheres to the mouse ischemic limbs ameliorated MSC survival, proliferation and paracrine effects under ischemic conditions. It also significantly accelerated angiogenesis, blood flow restoration, and skeletal muscle regeneration without provoking tissue inflammation. The above results demonstrate that the developed microspheres have potential to augment cell survival in ischemic tissues, and promote ischemic tissue regeneration in a safer and more efficient manner.

10.
Genome Biol ; 22(1): 16, 2021 Jan 05.
Artigo em Inglês | MEDLINE | ID: mdl-33402206

RESUMO

BACKGROUND: The folding of proteins is challenging in the highly crowded and sticky environment of a cell. Regulation of translation elongation may play a crucial role in ensuring the correct folding of proteins. Much of our knowledge regarding translation elongation comes from the sequencing of mRNA fragments protected by single ribosomes by ribo-seq. However, larger protected mRNA fragments have been observed, suggesting the existence of an alternative and previously hidden layer of regulation. RESULTS: In this study, we performed disome-seq to sequence mRNA fragments protected by two stacked ribosomes, a product of translational pauses during which the 5'-elongating ribosome collides with the 3'-paused one. We detected widespread ribosome collisions that are related to slow ribosome release when stop codons are at the A-site, slow peptide bond formation from proline, glycine, asparagine, and cysteine when they are at the P-site, and slow leaving of polylysine from the exit tunnel of ribosomes. The structure of disomes obtained by cryo-electron microscopy suggests a different conformation from the substrate of the ribosome-associated protein quality control pathway. Collisions occurred more frequently in the gap regions between α-helices, where a translational pause can prevent the folding interference from the downstream peptides. Paused or collided ribosomes are associated with specific chaperones, which can aid in the cotranslational folding of the nascent peptides. CONCLUSIONS: Therefore, cells use regulated ribosome collisions to ensure protein homeostasis.

11.
Neuromolecular Med ; 2021 Jan 03.
Artigo em Inglês | MEDLINE | ID: mdl-33389570

RESUMO

Glioma ranks first among the aggressive brain tumors all over the world. LncRNA LINC00689 has been confirmed to play key roles in the progression of cancers, and LINC00689 was upregulated in glioma. However, the biological function of LINC00689 in glioma is unclear. qRT-PCR was applied to detect the expressions of LINC00689 and miR-526b-3p in glioma cells. Dual-luciferase report was performed to examine the relation among LINC00689, miR-526b-3p, and insulin-like growth factor 2 mRNA-binding protein 1 (IGF2BP1). Then, the growth, migration, and invasion of glioma cells were detected by colony formation, flow cytometry, and transwell assay, respectively. The expressions of p21, cleaved caspase 3, and MAPK signaling-related proteins in glioma cells were tested by western blotting. Finally, xenograft mice model was established to detect the effect of LINC00689 on tumor growth of glioma in vivo. LINC00689 was upregulated in glioma cells, while miR-526b-3p was downregulated. In addition, LINC00689 bound to miR-526b-3p, and IGFBP1 was targeted by miR-526b-3p. Moreover, LINC00689 knockdown or upregulation of miR-526b-3p inhibited the proliferation of glioma cells and induced the apoptosis. Consistently, the migration and invasion of glioma cells were notably reduced by LINC00689 shRNA/miR-526-3p mimics. miR-526b-3p inhibitor or IGF2BP1 upregulation could reverse the effect of LINC00689 knockdown or miR-526b-3p mimics. Finally, knockdown of LINC00689 inhibited the tumor growth of glioma in vivo through regulating miR-526b-3p/IGF2BP1/MAPK axis. In conclusion, silencing of LINC00689 could inhibit the tumorigenesis of glioma via mediation of miR-526b-3p/IGF2BP1 axis. LINC00689 may serve as a new target for the treatment of glioma.

12.
Nanoscale ; 2021 Jan 13.
Artigo em Inglês | MEDLINE | ID: mdl-33439197

RESUMO

Micellar and vesicular structures capable of sensing and reporting the chemical environment as well as facilely introducing user-defined functions make a vital contribution to constructing versatile compartmentalized systems. Herein, by combining poly(ionic liquid)-based photonic spheres and an etching-ion exchange strategy we fabricate micellar and vesicular photonic compartments that can not only mimic the structure and function of conventional micelles and vesicles, but also sense and report the chemical environment as well as introducing user-defined functions. Photonic composite spheres composed of a SiO2 template and poly(ionic liquid) are employed to selectively etch outer-shell SiO2 followed by ion exchange and removal of the residual SiO2 to afford micellar photonic compartments (MPCs). The MPCs can selectively absorb solvents from the oil/water mixtures together with sensing and reporting the adsorbed solvents by the self-reporting optical signal associated with the uniform porous structure of photonic spheres. Vesicular photonic compartments (VPCs) are fabricated via selective infiltration and polymerization of ionic liquids followed by etching of the SiO2 template. Subsequent ion exchange introduces desirable functions to the VPCs. Furthermore, we demonstrate that the thickness and the anisotropic functions of VPCs can be facilely modulated. Overall, we anticipate that the micellar and vesicular photonic compartments with self-reporting optical signals and user-defined functions could serve as novel platforms towards multifunctional compartmentalized systems.

13.
Nat Commun ; 12(1): 678, 2021 01 29.
Artigo em Inglês | MEDLINE | ID: mdl-33514753

RESUMO

Reprogramming complex cellular metabolism requires simultaneous regulation of multigene expression. Ex-situ cloning-based methods are commonly used, but the target gene number and combinatorial library size are severely limited by cloning and transformation efficiencies. In-situ methods such as multiplex automated genome engineering (MAGE) depends on high-efficiency transformation and incorporation of heterologous DNA donors, which are limited to few microorganisms. Here, we describe a Base Editor-Targeted and Template-free Expression Regulation (BETTER) method for simultaneously diversifying multigene expression. BETTER repurposes CRISPR-guided base editors and in-situ generates large numbers of genetic combinations of diverse ribosome binding sites, 5' untranslated regions, or promoters, without library construction, transformation, and incorporation of DNA donors. We apply BETTER to simultaneously regulate expression of up to ten genes in industrial and model microorganisms Corynebacterium glutamicum and Bacillus subtilis. Variants with improved xylose catabolism, glycerol catabolism, or lycopene biosynthesis are respectively obtained. This technology will be useful for large-scale fine-tuning of multigene expression in both genetically tractable and intractable microorganisms.


Assuntos
Edição de Genes/métodos , Microbiologia Industrial/métodos , Engenharia Metabólica/métodos , Família Multigênica/genética , Bacillus subtilis/genética , Bacillus subtilis/metabolismo , Proteínas de Bactérias/genética , Proteínas de Bactérias/metabolismo , Sistemas CRISPR-Cas/genética , Corynebacterium glutamicum/genética , Corynebacterium glutamicum/metabolismo , DNA Bacteriano/genética , Escherichia coli/genética , Genes Bacterianos/genética , Glicerol/metabolismo , Licopeno/metabolismo , Redes e Vias Metabólicas/genética , Transformação Bacteriana , Xilose/metabolismo
14.
Nat Commun ; 12(1): 339, 2021 01 12.
Artigo em Inglês | MEDLINE | ID: mdl-33436626

RESUMO

Tuberous sclerosis complex (TSC) integrates upstream stimuli and regulates cell growth by controlling the activity of mTORC1. TSC complex functions as a GTPase-activating protein (GAP) towards small GTPase Rheb and inhibits Rheb-mediated activation of mTORC1. Mutations in TSC genes cause tuberous sclerosis. In this study, the near-atomic resolution structure of human TSC complex reveals an arch-shaped architecture, with a 2:2:1 stoichiometry of TSC1, TSC2, and TBC1D7. This asymmetric complex consists of two interweaved TSC1 coiled-coil and one TBC1D7 that spans over the tail-to-tail TSC2 dimer. The two TSC2 GAP domains are symmetrically cradled within the core module formed by TSC2 dimerization domain and central coiled-coil of TSC1. Structural and biochemical analyses reveal TSC2 GAP-Rheb complimentary interactions and suggest a catalytic mechanism, by which an asparagine thumb (N1643) stabilizes γ-phosphate of GTP and accelerate GTP hydrolysis of Rheb. Our study reveals mechanisms of TSC complex assembly and GAP activity.


Assuntos
Proteínas Ativadoras de GTPase/metabolismo , Proteína Enriquecida em Homólogo de Ras do Encéfalo/metabolismo , Proteína 1 do Complexo Esclerose Tuberosa/química , Proteína 1 do Complexo Esclerose Tuberosa/metabolismo , Proteína 2 do Complexo Esclerose Tuberosa/química , Proteína 2 do Complexo Esclerose Tuberosa/metabolismo , Biocatálise , Células HEK293 , Humanos , Modelos Moleculares , Ligação Proteica , Domínios Proteicos , Multimerização Proteica , Proteína 1 do Complexo Esclerose Tuberosa/ultraestrutura , Proteína 2 do Complexo Esclerose Tuberosa/ultraestrutura
15.
FEBS Open Bio ; 11(3): 598-621, 2021 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-33350590

RESUMO

Candida albicans is a harmless commensal resident in the human gut and a prevalent opportunistic pathogen. A key part of its commensalism and pathogenesis is its ability to switch between different morphological forms, including white-to-opaque switching. The Wor1 protein was previously identified as a master regulator of white-to-opaque switching in mating type locus (MTL) homozygous cells. The mechanisms by which the dark color of the opaque colonies is controlled and the pimpled surface of opaque cells is formed remain unknown. Candida albicans produces melanin pigment in vitro and during infection. However, the molecular mechanism underlying the regulation of melanin production is unclear. In this study, we demonstrated that ferroxidases (Fets) function as pigment multicopper oxidases and regulate the production of dark-pigmented melanin in opaque cells. The FET genes presented distinct regulation patterns in response to different extracellular stimuli. In YPD (1% yeast extract, 2% peptone and 2% dextrose)-rich medium, four of the five FET genes were up-regulated by Wor1, especially at the human body temperature of 37 °C. In minimal medium with low ammonium concentrations, all five FET genes were up-regulated by Wor1. However, at high ammonium concentrations, some FET genes were down-regulated by Wor1. Wor1-up-regulated Fets contributed to dark pigment formation in opaque colonies, but not to the elongated shape of these opaque cells. Increased melanin externalization was associated with the pimpled surface of the opaque cells. Melanized C. albicans cells were more resistant to fungal clearance. Deletion of the five FET genes completely blocked melanin production in opaque cells and resulted in the generation of white elongated 'opaque' cells. In addition, the up-regulated Fets are important for defense against oxidant attacks. The functional diversity of Fets may reflect the multiple strategies of C. albicans to rapidly adapt to diverse host niches.

16.
Int J Mol Sci ; 21(23)2020 Nov 26.
Artigo em Inglês | MEDLINE | ID: mdl-33255884

RESUMO

Inflammation is the main pathophysiology of dry eye, characterized by tear film instability and hyperosmolarity. The aim of this study was to investigate the association of inflammation and cellular autophagy using an in vitro dry eye model with primary cultured human corneal epithelial cells (HCECs). Primary HCECs cultured with fresh limbal explants from donors were switched to a hyperosmotic medium (450 mOsM) by adding sodium chloride into the culture medium. We observed the stimulated inflammatory mediators, TNF-α, IL-1ß, IL-6 and IL-8, as well as the increased expression of autophagy related genes, Ulk1, Beclin1, Atg5 and LC3B, as evaluated by RT-qPCR and ELISA. The immunofluorescent staining of LC3B and Western blotting revealed the activated autophagosome formation and autophagic flux, as evidenced by the increased LC3B autophagic cells with activated Beclin1, Atg5, Atg7 and LC3B proteins, and the decreased levels of P62 protein in HCECs. Interestingly, the autophagy activation was later at 24 h than inflammation induced at 4 h in HCECs exposed to 450 mOsM. Furthermore, application of rapamycin enhanced autophagy activation also reduced the inflammatory mediators and restored cell viability in HCECs exposed to the hyperosmotic medium. Our findings for the first time demonstrate that the autophagy activation is a late phase response to hyperosmotic stress, and is enhanced by rapamycin, which protects HCECs by suppressing inflammation and promoting cells survival, suggesting a new therapeutic potential to treat dry eye diseases.

17.
ACS Infect Dis ; 2020 Dec 08.
Artigo em Inglês | MEDLINE | ID: mdl-33291867

RESUMO

Mutations in KPC-2 and KPC-3 ß-lactamase can confer resistance to the ß-lactam/ß-lactamase inhibitor antibacterial intravenous drug combination ceftazidime-avibactam, introduced in 2015. Avibactam was the first of the diazabicyclooctane class of non-ß-lactam ß-lactamase inhibitors to be approved for clinical use. The orally bioavailable prodrug ETX0282 of the diazabicyclooctane ß-lactamase inhibitor ETX1317 is in clinical development in combination with the oral ß-lactam prodrug cefpodoxime proxetil for use against complicated urinary tract infections. We investigated the effects of 3 ceftazidime-avibactam resistance mutations in KPC-3 (V240G, D179Y, and D179Y/T243M) on the ability of ETX1317 to overcome KPC-3-induced cefpodoxime resistance. Isogenic Escherichia coli strains, each expressing the wild-type or a mutant KPC-3 at similar levels, retained susceptibility to cefpodoxime-ETX1317 (1:2) with essentially identical minimal inhibitory concentrations of 0.125-0.25 µg/mL cefpodoxime. The KPC-3 mutations had little or no effect on the kinact/Ki values for inhibition by each of 3 diazabicyclooctanes: avibactam, durlobactam (ETX2514), and ETX1317. The KM values for hydrolysis of cefpodoxime were similar for all 4 variants, but the kcat values of the D179Y and D179Y/T243M variants were much lower than those of the wild-type and V240G mutant enzymes. All 4 KPC-3 variants formed stable, reversibly covalent complexes with ETX1317, but dissociation of ETX1317 was much slower from the D179Y and D179Y/T243M mutants than from the wild-type and V240G mutant enzymes. Thus, the KPC-3 variants examined here that cause resistance to ceftazidime-avibactam do not cause resistance to cefpodoxime-ETX1317.

18.
Cell Discov ; 6(1): 88, 2020 Nov 24.
Artigo em Inglês | MEDLINE | ID: mdl-33298899

RESUMO

The function of the origin recognition complex (ORC) in DNA replication is highly conserved in recognizing and marking the initiation sites. The detailed molecular mechanisms by which human ORC is reconfigured into a state competent for origin association remain largely unknown. Here, we present structural characterizations of human ORC1-5 and ORC2-5 assemblies. ORC2-5 exhibits a tightly autoinhibited conformation with the winged-helix domain of ORC2 completely blocking the central DNA-binding channel. The binding of ORC1 partially relieves the autoinhibitory effect of ORC2-5 through remodeling ORC2-WHD, which makes ORC2-WHD away from the central channel creating a still autoinhibited but more dynamic structure. In particular, the AAA+ domain of ORC1 is highly flexible to sample a variety of conformations from inactive to potentially active states. These results provide insights into the detailed mechanisms regulating the autoinhibition of human ORC and its subsequent activation for DNA binding.

19.
ACS Infect Dis ; 2020 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-33300345

RESUMO

ETX0282 is an orally bioavailable prodrug of the diazabicyclooctane serine ß-lactamase inhibitor ETX1317. The combination of ETX0282 with cefpodoxime proxetil is in clinical trials as an oral therapy for complicated urinary tract infections caused by Enterobacterales. Earlier diazabicyclooctane ß-lactamase inhibitors, such as avibactam and durlobactam, contain a sulfate moiety as the essential anionic group and are administered intravenously. In contrast, ETX1317 contains a fluoroacetate moiety, which is esterified with an isopropyl group in ETX0282 to provide high oral bioavailability. Previous studies of avibactam and durlobactam showed that covalent inhibition of certain ß-lactamases is reversible due to the ability of the ring-opened inhibitors to recyclize and dissociate in their original form. We investigated the interaction of ETX1317 with several ß-lactamases commonly found in relevant bacterial pathogens, including CTX-M-15, KPC-2, SHV-5, and TEM-1 from Ambler Class A; Pseudomonas aeruginosa AmpC and Enterobacter cloacae P99 from Class C, and OXA-48 from Class D. The second-order rate constants for inhibition (kinact/Ki) of these enzymes show that ETX1317 is intermediate in potency between durlobactam and avibactam. The partition ratios were all approximately 1, indicating that the inhibitor is not also a substrate of these enzymes. The rate constants for dissociation of the covalent complex (koff) were similar to those for durlobactam and avibactam. Acylation exchange experiments demonstrated that ETX1317 dissociated in its original form. No loss of mass from the inhibitor was observed in the covalent inhibitor-enzyme complexes.

20.
Artigo em Inglês | MEDLINE | ID: mdl-33326866

RESUMO

The distal end of parotid duct is often inevitably resected en-block with the buccal mucosa cancer to obtain safety margin and prevent local recurrence. Ligation of duct frequently causes complications like cheek swelling, fistula and gland function loss. The authors describe a novel procedure of combined use of autologous vein graft and vascular coupler to reconstruct the parotid duct defect for buccal mucosa cancer patients who undergo radical neck dissection and free-flap reconstruction, no lumen-stent or cannula is needed. Case examples are shown to illustrate the operative details and different outcomes for two kinds of orifice site choices. Key factors for success include the proper use of Coupler device, right choice of new orifice location and maintenance of lumen patency. For buccal mucosa cancer patients, this novel method for parotid duct reconstruction could effectively reduce postoperative complications and preserve the parotid gland function.

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